The intracelluar Na ion activity (aiNa) and the tension (T) of sheep cardiac Purkinje fibers were simultaneously measurd employing recessed-tip Na ion-selectve glass microelectrodes and a mechano-electric transducer. The aiNa of 6.4 plus or minus 1.6 mM (mean plus or minus S.D., n equals 56) was obtained in fibers perfused with normal Tyrode's solution. The changes in aiNa and T were measured during and after the exposure of fibers to a cardiac glycoside, dihydro-ouabain (DHO) in concentrations between 5 x 10 to the minus 8th power M and 10 to the minus 5th power M. The exposure time to DHO was 15 min. Both aiNa and T did not change in fibers exposed to 5 x 10 to the minus 8th power M DHO, and the threshold concentration for the effect of DHO appeared to be around 10 minus to the 7th power M. The increase in DHO concentration above the threshold resulted in progressively greater increased in both aiNa and T. The increased in aiNa were linearly related to those in T/Tc (the ratio of increased tension to control tension). During onset of DHO effects, the time course of aiNa increase was concomittant with that of T increase. The recoveries of aiNa and T were variable and slow, being dependent on the DHO concentration. In those fbers which recovered from the effects of DHO, the time course of aiNa recovery was similar to that of T recovery. In fibers exposed to DHO of 5 x 10 to the minus 6th power M or greater, the apparent toxic effects were observed in both action potential and contraction after an initial increase in T. The fibers manifesting the apparent toxic effects had the aiNa of about 30 mM or greater. The results in this study indicate that the increase in aiNa is coincided with the positive inotropic action of the cardiac glycoside.